P409: BCL-2/BCL-XL INHIBITION INDUCES APOPTOSIS AND CIRCUMVENTS VENETOCLAX RESISTANCE IN TP53-MUTATED ACUTE MYELOID LEUKEMIA
Authors: Ida Vänttinen, Sari Kytölä, Olli Dufva, Monica Hellesøy, Stein-Erik Gullaksen, Tanja Ruokoranta, Oscar Brück, Sanna Siitonen, Pia Ettala, Marja Pyörälä, Johanna Rimpilainen, Timo Siitonen, Bjorn Tore Gjertsen, Caroline Heckman, Heikki Kuusanmäki, Mika Kontro
Journal: HemaSphere (2023)

Background: The tumor suppressor TP53 gene is a fundamental regulator of cellular processes with functions ranging from DNA repair to induction of apoptosis. TP53 alterations occur in about 5-15% of acute myeloid leukemia (AML) cases and are associated with particularly poor treatment outcomes. Recent reports have demonstrated venetoclax-based therapy to be ineffective in TP53mut AML. Dysfunctional TP53 might prevent the apoptosis-inducing effect of venetoclax by impairing the pro-apoptotic BAX/BAK activation required to trigger cell death. Identifying alternative druggable targets to overcome venetoclax resistance and to ultimately induce apoptosis in TP53mut AML is therefore urgently needed. Aims: To analyze the genetic, proteomic and ex vivo BH3 mimetic drug screening profiles of 109 AML patients enrolled in the VenEx trial (NCT04267081) to identify venetoclax resistance mechanisms in TP53mut AML. Methods: 109 AML samples’ blast-specific ex vivo responses to venetoclax (BCL-2 inhibitor), navitoclax (BCL-2/BCL-XL inhibitor), A-1331852 (BCL-XL inhibitor) and S-63845 (MCL-1 inhibitor) were profiled by flow cytometry. Mutations in the respective patients were analyzed using targeted panel sequencing. Expression levels of 19 intracellular proteins including BCL-2 family members (BCL-2, BCL-XL, MCL-1, BAX, BAK, BIM, BAD, PUMA) of the CD34+ blasts were quantified by CyTOF mass cytometry (n=36/109). BCL-2 protein expression of the blasts of another independent AML cohort (n=53) was analyzed by flow cytometry. Genetic and proteomic associations with the ex vivo drug sensitivities were computed using the Wilcoxon rank sum test and Spearman’s correlation, respectively. Results: In the computational comparison analysis of VenEx trial participants’ genetic aberrations and BH3 mimetic ex vivo responses, TP53 mutation (p=0.005) and complex karyotype (p=0.003) were the strongest predictors of venetoclax resistance, while IDH2 (p=0.006) and SRSF2 (p=0.007) mutations predicted favorable responses (Figure A). In contrast, TP53mut patients (n=12/109) had increased sensitivity to A-1331852 (Figure A), and the dual BCL-2/BCL-XL inhibitor navitoclax was highly effective in TP53mut blasts when compared to single agents venetoclax and A-1331852 (Figure B). MCL-1 inhibitor (S-63845) efficacy, however, was not associated with TP53mut or complex karyotypic changes. Aligned with decreased venetoclax sensitivity of TP53mut blasts, the CyTOF analysis revealed that BCL-2 protein expression was significantly lower in those samples compared to TP53 wild-type counterparts (p=0.03), but no significant differences in BCL-XL, BAX, BAK and PUMA levels were observed. To further validate the BCL-2 protein expression pattern, we investigated another independent AML cohort, where a similar trend of decreased expression of BCL-2 in TP53mut blasts was detected (p=0.06, Figure C). Summary/Conclusion: This study showed that AML patients harboring TP53 mutations have reduced ex vivo sensitivity to venetoclax, which might be associated with decreased BCL-2 protein expression. In contrast, our BH3 mimetic drug screening results demonstrated that the dual inhibition of BCL-2 and BCL-XL by navitoclax is capable of inducing apoptosis in myeloid blast cells regardless of TP53 mutation status. However, larger patient cohorts and more extensive functional analysis of the anti/pro-apoptotic dependencies in this patient group is warranted.Keywords: TP53, Drug resistance, Acute myeloid leukemia, Venetoclax